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PubMed1979

Ultrastructure of basophilic leukocytes and mast cells in normal and cutaneous basophil hypersensitivity-reacted guinea pig dermis.

Basophilic leukocytes and mast cells in guinea pig dermis in normal and cutaneous basophil hypersensitivity (CBH) reaction were examined by light and electron microscopy. Basophils were rare in the normal dermis and predominantly revealed in the CBH-reacted skin. Some infiltrating basophils of the reacted displayed an immediate attachment to mast cells. Cytoplasmic continuities were partially seen between them. They were most plentiful at 48 h after phytohemagglutinin injection and decreased in number thereafter. The basophils contained three types of granule. The vast majority of granules were Type I granules, basophil-specific granules. Type II granules were less frequently encountered and resembled a mast cell granule in the fine structure. Type III granules were scant and small-cored vesicles.

Animals↗
PubMed2006

When is a mouse basophil not a basophil?

The identification and characterization of mouse basophils have historically been hampered by the extreme rarity of this cell type. Virtually no photomicrographs of hematologically stained (eg, Wright-Giemsa) examples of mouse basophils exist in the literature. However, 4 recent studies in the past 2 years have used flow cytometry and a defined set of cell-surface markers to identify and subsequently isolate mouse "basophils," including the publication of stained cytospin preparations of these cells. Surprisingly, a reevaluation of the data from all 4 of the studies revealed several issues of concern that suggest that the cells under study are not necessarily basophils. Nonetheless, we propose that these studies do provide the foundation for a reevaluation of the defining characteristics of a basophil and/or provide support for the provocative conclusion that a new previously overlooked leukocyte subtype has been identified. The purpose of this commentary is to revisit these previously published studies, highlight the relevant issues, and provide a different perspective in the hope of developing a consensus within the research community as to the true identity of the "basophils" described in these studies.

Animals↗
PubMed2001

Characteristics of the free cytosolic calcium timelag following IgE-mediated stimulation of human basophils: significance for the nonreleasing basophil phenotype.

These studies examine characteristics of the quiescent period (timelag) of the free cytosolic calcium ([Ca++]i) elevation that follows stimulation of human basophils through the IgE receptor. Previous studies established that the [Ca++]i timelag was sensitive to the rate of ligand binding, but little else is known about this response characteristic. The [Ca++]i timelag could be lengthened using antigenic stimulation that is rapid but only weakly induces secretion: tenfold differences in the "strength" of the stimulus, as assessed by histamine release, are associated with threefold differences in the timelag. Inhibiting p53/56lyn kinase with low concentrations of the specific inhibitor, PP1, lengthened the [Ca++]i timelag dramatically. PP1 was also found to delay the onset of syk phosphorylation and histamine release. Staurosporine and genistein, which are known to inhibit early tyrosine kinases, had, at best, only modest effects on the [Ca++]i timelag. Specific inhibitors of protein kinase C (PKC) had no effect on the [Ca++]i timelag, and direct activation of PKC with PMA had only very modest effects on the timelag. Contrary to expectations, basophils with the so-called nonreleasing phenotype demonstrated an IgE-mediated [Ca++]i response at the single-cell level. However, the length of [Ca++]i timelag in nonreleasing basophils was threefold longer than normally found in releasing basophils. Furthermore, the [Ca++]i response was significantly more asynchronous than in releasing basophils and lacking in a sustained [Ca++]i elevation. These studies indicate that the [Ca++]i timelag following stimulation through the IgE receptor is sensitive to inhibition of lyn kinase but not other agents that have been demonstrated to inhibit early tyrosine kinases previously. However, only one characteristic of the [Ca++]i response phenotype of nonreleasing basophils--the [Ca++]i timelag but not the absence of a sustained [Ca++]i elevation--could be mimicked by inhibition of lyn kinase with PP1.

Adult↗
PubMed1989

[Studies on human basophil and eosinophil colonies in human blood mononuclear cell culture: presence of common basophil-eosinophil progenitors].

Human peripheral blood mononuclear cells (PBMC) obtained from healthy volunteers gave rise to three types of colonies that could be distinguished by their unique morphological characteristics; 5.1 +/- 0.9 (mean +/- SE) loose colonies of small cells, 2.1 +/- 0.9 packed colonies of larger cells, and 1.6 +/- 0.7 mixed colonies, were formed when PBMC (2 x 10(5) cells/dish) from 10 healthy volunteers were cultured in the medium containing methylcellulose. Cytochemical analysis with Astra blue-eosin dual staining revealed that all types of colonies consisted of various proportions of basophils, eosinophils, and hybrid (eosinophilic and basophilic) granulocytes which contained both of granules. These hybrid granulocytes were also identified by the ultrastructural features of two kinds of the granules. Relationships between cell numbers added to culture and formed colony numbers indicated colony of the cells to form the colonies. The colonies formed from untreated patients with chronic myelogenous leukemia (CML) during chronic phase were sevenfold of those from healthy volunteers. The colonies formed from treated patients with CML were normal in number. The number was 40 times greater in culture from a patient with basophilic crisis and a patient with myeloid crisis than normal, whereas that from a patient with lymphoid crisis were within normal limit. The number of the colonies from PBMC of patients with eosinophilia were in normal range, whereas those from bone marrow were six times or more than those from PBMC. These findings suggest that PBMC contains common basophil-eosinophil progenitors, and the culture used in this study in considered to be useful in the examination of basophil and eosinophil production from PBMC. Further studies using more purified cell population and other sources of colony stimulating factors such as interleukin-3 (IL-3), IL-4, IL-5, and granulocyte-macrophage colony stimulating factor should be carried out in order to clarify the significance of hybrid granulocytes in basophil and eosinophil proliferation and differentiation.

Adult↗
PubMed1980

Generation and release of platelet-activating factor (PAF) from enriched preparations of rabbit basophils; failure of human basophils to release PAF.

Rabbit mononuclear cells containing up to 20% basophils and uncontaminated by neutrophils release PAF when stimulated with goat antiserum to rabbit IgE. The amount of PAF detected was a function of basophil concentration but decreased on a per cell basis at high basophil or high total cell concentrations. Calcium ionophore A23187, but not protein A, C5a, or FMLP, initiated rabbit basophil degranulation and PAF release. By contrast, extensive studies using a variety of human leukocyte preparations failed to demonstrate the release of significant levels of PAF from human basophils by IgE-dependent or -independent mechanisms. These results suggest that cells other than the peripheral blood basophil (e.g., the neutrophil) may act as the primary site of PAF production in man.

Animals↗
PubMed1984

Inhibition by somatostatin of the release of mediators from human basophils and rat leukemic basophils.

The release of mediators from unpurified human basophils challenged with anti-human myeloma IgE serum was suppressed significantly by somatostatin at respective concentrations of 3 x 10(-13) M to 10(-9) M for histamine and 10(-13) M to 10(-11) M for leukotriene D4, which had no effect on release elicited by ionophore A23187. A direct effect of picomole-nanomole concentrations of somatostatin on basophils predominated, as shown by the significant inhibition of immunologically stimulated mediator release from monocyte-free human basophils of 15 to 32% purity and from rat basophil leukemia cells. The dependence of basophil inhibition on the conformation of somatostatin was suggested by the lower potency of reduced and alkylated somatostatin and the lack of inhibitory activity of [D-Trp8] somatostatin. Somatostatin thus may mediate suppressive effects of sensory nerves in some basophil-dependent hypersensitivity reactions.

Animals↗
PubMed1982

Dissociation between cutaneous basophil hypersensitivity and basophil sensitization.

We investigated the antigen-specific sensitization of basophils in the peripheral circulation of guinea pigs using an in vitro histamine release assay. Animals were primed for cutaneous basophil hypersensitivity (CBH) by intradermal injection of either keyhole limpet hemocyanin (KLH) or ovalbumin (OA) in saline. Basophils isolated from these animals 7 days after sensitization did not release histamine to the specific antigen in vitro. On day 9, however, basophils did release histamine when challenged with the antigen. These results were in contrast to 24-hr skin tests that showed a typical CBH profile both on day 7 and day 9. When animals were immunized via the footpad by emulsification of antigen in incomplete Freund's adjuvant, basophil sensitization (i.e., histamine release) either occurred earlier (day 7, KLH) or at lower antigen concentrations (OA). Thus, dissociation of in vivo CBH and in vitro basophil sensitization was demonstrated. The lack of correlation was only seen early after immunization and depended on the route of immunization used. An additional finding was a transient blood basophilia that also was dependent on the route of immunization, the antigen used, and the time post-sensitization. Our results are consistent with the hypothesis that at early times post-immunization, CBH is mediated by T cells and not via homocytotropic antibodies, but that at later times such antibodies may also play a role in the elicitation of CBH reactions.

Animals↗
PubMed1996

Basophilic deposits on the cupula: preliminary findings describing the problems involved in studies regarding the incidence of basophilic deposits on the cupula.

In this study, the possibility of whether basophilic deposits adhered to the cupulas in the semicircular canals was investigated histologically. Results indicated that basophilic deposits were present in all three cupulas of the semicircular canals. The overall incidence of basophilic deposits in the superior, lateral and posterior semicircular canal cupulas was 26%, 41% and 37%, respectively. The incidence of basophilic deposits bound to the cupulas increased with age. The possible origin of these basophilic deposits on the cupulas and the increased incidence of basophilic deposits with increasing age are discussed.

Adult↗
PubMed1991

Regulation of human basophil activation. I. Dissociation of cationic dye binding from histamine release in activated human basophils.

Human basophil activation was demonstrated by histamine release (HR) and by the decrease of the toluidine blue-positive basophils (TB+). In four experimental systems, TB+ number decreased in the absence of HR (1) in basophils from atopic subjects stimulated by allergen concentrations below the threshold for HR, (2) in basophils sensitized by anti-2,4-dinitrophenyl IgE stimulated by noncovalently linked 2,4-dinitrobenzene sulfonic acid-human serum albumin (also, the threshold for decrease of TB+ required lower concentrations of sensitizing anti-2,4-dinitrophenyl IgE than for HR), (3) in low Ca++ medium, and (4) in the presence of the Na+/H+ exchanger, monensin. These results suggest that (1) there is a lower threshold for TB+ decrease than for HR in allergen concentration, number of membrane IgE molecules, and number of IgE cross-linkings; moreover, external Ca++ requirement is lower for decrease of TB+ than for HR and (2) TB+ decrease reflects either granule exocytosis or, in the absence of HR, biochemical changes (most probably cation exchanges) altering the interaction of the basic dye with the granules. Thus, monitoring decrease in TB+ allows detection of basophil activation in the absence of HR.

Basophils↗
PubMed1981

Flow-cytometric analysis of human basophil degranulation. I. Quantification of human basophils and their degranulation by flow-cytometry.

Quantification of human basophils and their degranulation were performed using a flow-cytometric system. It was shown that the number of basophils among total leukocytes, before and after the effect of degranulating agents, can be obtained rapidly and reproducibly, following alcian blue staining. Dose-response curves of degranulation by anti-IgE and anti-IgG4 were studied, and it was shown that these antibodies can induce basophil degranulation in a Ca++ dependent manner. It was also shown that degranulation is completed within 10 min. Values obtained by flow-cytometry and microscopic evaluation, as well as comparison made between percent basophil degranulation and percent histamine release, agreed well with each other in our system. Flow-cytometric analysis of human basophil degranulation provides a new reliable method to assess in vitro the morphological basis of immediate hypersensitivity.

Antibodies, Anti-Idiotypic↗
PubMed1975

Specificity of basophils and lymphocytes in cutaneous basophil hypersensitivity.

Using a rosetting technique, it was found that the vast majority of basophils circulating in the blood or accumulating in the skin reactions of guinea pigs primed for cutaneous basophil hypersensitivity (CBH) lacked demonstrable specificity for sensitizing antigen, whether sheep erythrocytes, a soluble protein, or tumor cells. By contrast, one-third of cells teased from late skin reactions formed specific rosettes as did nearly 80% of circulating basophils in animals receiving repeated doses of whole sheep blood. Unreactive basophils teased from CBH reactions readily acquired rosetting capacity on exposure to immune serum. With regard to lymphocyte (and hence reaction) specificity, both CBH and classic delayed hypersensitivity (DH) reactions exhibited a high degree of carrier specificity when dinitrophenyl-conjugates were used. Thus, in the hapten-carrier combinations examined thus far, the antigen skin test requirements for both CBH and DH have been identical and are those required for inducing an active lymphocyte response. These findings indicate that control mechanisms other than homocytotropic antibodies must be sought to explain the accumulation and behavior of basophils in CBH reactions and, coupled with other data, suggest that lymphocytes and/or their products are likely candidates for this role.

Animals↗
PubMed1982

Use of automatic counting of basophil leukocytes: correlation of basophil degranulation with histamine release.

The sensitivity of leukocytes from allergic donors towards specific allergen was studied using the basophil degranulation test. In this test, basophil leukocytes were counted with the Technicon Hemalog D. A good correlation was found between the results of the basophil degranulation test and the results of the histamine release test on leukocytes of the same patient. The basophil degranulation test, using the Hemalog D, was found to be a simple method for in vitro assessment of the sensitivity of allergic patients to specific allergen.

Allergens↗
PubMed1994

Regulation of human basophil activation; the role of Na+ and Ca2+ in IL-3-induced potentiation of IgE-mediated histamine release from human basophils.

The release of mediators from human basophils is strongly enhanced by IL-3. However, the signalling pathways of IL-3 are poorly defined in these cells. Since external Ca2+ and Na+ play important regulating roles in histamine release, the possibility that these cations could be involved in the potentiation by IL-3 of the anti-IgE-induced histamine release from human basophils was considered, and it was observed that: (i) IL-3 dramatically decreased the external Ca2+ requirement for IgE-mediated histamine release. However, histamine release from IL-3-treated basophils became only partially independent of external Ca2+, since addition of EGTA in the external medium abolished the effect of IL-3; (ii) decreasing Na+ influx by lowering external Na+ concentration in isosmotic medium inhibited the potentiating effect of IL-3 on IgE-mediated release; (iii) amiloride, an inhibitor of Na+/Ca2+ and Na+/H+ exchanges, and its derivative, benzamil, more specific for Na+/Ca2+ exchanges, inhibited the release potentiated by IL-3. In contrast, the amiloride derivative 5-(N,N-dimethyl)-amiloride, more specific for Na+/H+ exchanges, slightly increased the IL-3-enhanced release. Thus, the decreased requirement for external Ca2+ and the dependence on external Na+, taken with the effect of the Na+/Ca2+ exchange inhibitors, suggest that Na+/Ca2+ exchanges are involved in the IL-3-induced enhancement of IgE-mediated human basophil histamine release.

Amiloride↗
PubMed1977

A simplified method for measuring basophil histamine release and blocking antibodies in hay fever patients. Basophil histamine content and cell preservation.

A simplified method for measuring basophil histamine release in grass pollen hay fever patients has been developed. Leukocytes were challenged in vitro with extracts of Phleum pratense (timothy) and the release of histamine was determined indirectly as the residual histamine in the cell sediment. Several steps to purify histamine thus became superfluous and histamine was directly conjugated with o-phthaldialdehyde to form a fluorophore. The simplified method showed a basophil histamine content which was in accordance with results obtained by more specific methods. No difference in basophil histamine content was found between normal and allergic persons. For the histamine liberation assay blood could be adequately preserved for transport for 48 h at room temperature by adding cell culture medium. Basophil histamine release technique allows evaluation of cell sensitivity for determination of the degree of allergy as well as the level of blocking antibodies.

Adult↗
PubMed1981

Basophil production. IV. Morphology of basophils in liquid culture.

Guinea pig bone marrow basophils were cultivated in vitro and morphological observations were made by light and electron microscopy. Maturation of basophils in vitro is supported by electron microscopic evidence for the development of a prominent Golgi zone with granule formation, nuclear chromatin changes, and specific periodicity of the intragranular matrix. Histochemical staining distinguishes these cells from other granulocytes, and mitotic basophils are recognizable on light microscopy. These methods allow more detailed study of the orderly growth and development of the basophil in a system where basophilopoiesis is actively taking place.

Animals↗
PubMed1986

Human basophil releasability. III. Genetic control of human basophil releasability.

Basophil releasability implies that, in addition to the surface density of IgE molecules, biochemical events determine the capacity to release chemical mediators in response to activating stimuli. We studied the IgE (anti-IgE)-mediated and non-IgE-mediated (f-met peptide and the Ca2+ ionophore A23187) releasability of human basophils obtained from 14 monozygotic (MZ) (ages 25.7 +/- 13.3 yr; mean +/- SDM) and 13 dizygotic (DZ) twin pairs (ages 20.4 +/- 9.9 yr). A significant intrapair correlation coefficient of the maximal percent of anti-IgE-induced histamine release was found in the MZ, whereas no significant correlation was found in the DZ. The mean intrapair variance of anti-IgE-induced histamine release in MZ (VMZ) and in DZ (VDZ) gave an F value equal to 3.84 (p less than 0.01) and a heritability (H) index of 0.74. Similar findings were obtained with respect to the sensitivity to a standard concentration (10(-1) micrograms/ml) of anti-IgE. No correlation between serum IgE level and anti-IgE-induced histamine release was found in either MZ or DZ. A significant intrapair correlation coefficient of f-met peptide-induced histamine release was found in both the MZ and the DZ. The difference between MZ and DZ was not significant. The VMZ and the VDZ of the f-met peptide-induced histamine release gave an F value of 1.52 (NS) and an H value of 0.34. The intrapair correlation coefficient of A23187-induced release was significant in MZ and not significant in DZ. The mean intrapair variance of A23187-induced histamine release gave an F value of 2.33 (NS) and an H index of 0.57. Similar findings were obtained by using suboptimal (3 X 10(-1) micrograms/ml) concentrations of A23187. There was no correlation between the sensitivity of basophils to release in response to anti-IgE and their response to f-met peptide or A23187, in either the MZ or the DZ. We conclude that the ability of basophils to respond to anti-IgE and A23187 is influenced by genetic factors.

Antibodies, Anti-Idiotypic↗
PubMed1996

The effect of okadaic acid on histamine release, cell morphology and phosphorylation in rat basophilic leukemia (RBL-2H3) cells, human basophils and rat peritoneal mast cells.

To study the involvement of serine/threonine phosphatase in the signal transduction of mast cells, we examined the effects of okadaic acid (OA), an inhibitor of type-1 and -2A phosphatase on histamine release, cell morphology, calcium influx and protein phosphorylation of rat basophilic leukemia (RBL-2H3) cells, human basophils and rat peritoneal mast cells. OA inhibited IgE-mediated histamine release from RBL-2H3 cells and human basophils dose-dependently. There was a remarkable enhancement of IgE-mediated histamine release when rat peritoneal mast cells were suboptimally challenged. OA induced a marked change of cell features, detached RBL-2H3 cells from plastic well and kept the 18- and 68-kD proteins phosphorylated. These findings show that phosphatase may play a role in the modulation of secretion in mast cells.

Animals↗